Among the most debilitating and costly human ailments are injuries and diseases of the nervous system. They affect millions of people in the US and represent a large part of the total national health care cost. Currently there are a limited number of available therapies, none of which restore function to injured neurons of the CNS. Research efforts from a variety of areas are offering real hope for improving the clinical outcome of these conditions. Numerous studies in animals and man strongly suggest that restorative therapies based on cell transplantation are feasible. However, a major challenge that remains is the reconstruction of damaged and diseased neural pathways. Toward this end, biomaterials have been examined as bridging devices to support directed nerve outgrowth from regenerating neurons. One of the fundamental challenges involves understanding how to engineer the surface of such biomaterial bridges. This revised Bioengineering Research Partnership (PAS- 00-006) project is driven by the central hypotheses that the local surface density, conformation and discrete spatial distribution of substrate molecules are sensed by a neuron's integrin receptors, translated in an intracellular molecular sequence that regulates integrin receptor expression and controls further axonal growth and also determines the overall readiness of the neuron to regenerate and establish connections. This partnership will bring together the expertise of a surface analytical chemist (Beebe) possessing a background in bond- rupture force measurements, surface modification, and state-of- the-art surface analytical techniques, with the expertise of a biophysics-oriented bioengineer (Hlady) possessing a background in protein-surface interactions, evanescent wave optics, time- resolved fluorescence spectroscopy, scanning probe microscopy and a complementary set of state-of-the-art surface analytical techniques and a neuroscience-oriented bioengineer (Tresco) possessing a background in tissue engineering, nervous system repair and biomaterials. The three specific aims are to: (1) create and fully characterize model substrates with a controlled pattern and surface density of laminin, fibronectin and related oligopeptides for neuron and astrocyte attachment and axonal growth studies in the two other Aims; (2) Study neurite outgrowth (dynamic bond strength, diffusivity, surface density and bond-rupture force) of dorsal root ganglion neurons on the model substrates, employing single- molecule techniques such as AFM bond-rupture measurements and fluorescence correlation spectroscopy; (3) Study the axonal growth of neurons (dynamic bond strength, diffusivity, surface density and bond-rupture force) on confluent astrocyte monolayers of different ages, using the same methodologies. Underpinning all studies are extensive quantitative surface characterization techniques. The advent and maturation of the methodologies used in single- molecule bond-rupture and single-molecule fluorescence measurements have allowed the direct and highly controlled measurement of protein-ligand, protein-protein, and protein- surface interactions on an individual, molecule-by-molecule basis. Similar ligand-receptor studies (i.e. binding-force measurements complemented with single-molecule fluorescence experiments) will be extended to the study of molecules involved in the guidance of neuron growth cones and axons. This molecular understanding of the balance between specific and non-specific ligand-receptor interactions will be supplemented with microscopy studies (fluorescence and atomic force) of axon development on ligand-modified substrates.